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The role of iron in Mycobacterium smegmatis biofilm formation: the exochelin siderophore is essential in limiting iron conditions for biofilm formation but not for planktonic growth.

Ojha A, Hatfull GF - Mol. Microbiol. (2007)

Bottom Line: In contrast, although the expression of mycobactin and iron ABC transport operons is highly upregulated during biofilm formation, mutants in these systems form normal biofilms in low-iron (2 microM) conditions.A close correlation between iron availability and matrix-associated fatty acids implies a possible metabolic role in the late stages of biofilm maturation, in addition to the early regulatory role.M. smegmatis surface motility is similarly dependent on iron availability, requiring both supplemental iron and the exochelin pathway to acquire it.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260, USA.

ABSTRACT
Many species of mycobacteria form structured biofilm communities at liquid-air interfaces and on solid surfaces. Full development of Mycobacterium smegmatis biofilms requires addition of supplemental iron above 1 microM ferrous sulphate, although addition of iron is not needed for planktonic growth. Microarray analysis of the M. smegmatis transcriptome shows that iron-responsive genes - especially those involved in siderophore synthesis and iron uptake - are strongly induced during biofilm formation reflecting a response to iron deprivation, even when 2 microM iron is present. The acquisition of iron under these conditions is specifically dependent on the exochelin synthesis and uptake pathways, and the strong defect of an iron-exochelin uptake mutant suggests a regulatory role of iron in the transition to biofilm growth. In contrast, although the expression of mycobactin and iron ABC transport operons is highly upregulated during biofilm formation, mutants in these systems form normal biofilms in low-iron (2 microM) conditions. A close correlation between iron availability and matrix-associated fatty acids implies a possible metabolic role in the late stages of biofilm maturation, in addition to the early regulatory role. M. smegmatis surface motility is similarly dependent on iron availability, requiring both supplemental iron and the exochelin pathway to acquire it.

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Schematic representations of transcriptome responses in 3 day biofilm, 4 day biofilm and stationary-phase conditions. A. Conditions corresponding to the 3 day biofilm (green circle), 4 day biofilm (blue circle) and stationary phase (red circle) are shown, with the numbers of induced genes (> 4-fold) shown in black and the number of repressed genes (> 4-fold) shown in red. The numbers of induced and regulated genes common to two or more conditions are included at the intersections of the three circles. A complete list of microarray data for genes in each category is included in Table S2. B. Induction of genes involved in iron acquisition during biofilm and stationary-phase growth. Twenty-nine genes arranged in nine operons (Msmeg0011–0014, Msmeg0015, Msmeg0016–0018, Msmeg2133–2135, Msmeg4502–4509, Msmeg4510, Msmeg5028, Msmeg6024–6026 and Msmeg6214–6216) putatively involved in iron acquisition show substantial levels of induction after 4 days of biofilm development. Arrows indicate the operon arrangements. Average fluorescence units are shown for the exponential planktonic (blue bars), 3 day biofilm (red bars), 4 day biofilm (yellow bars) and stationary phase (green bars). Fluorescence values for the exponential planktonic sample are the average of 12 normalized slides, and for the other three experimental conditions, are the average from three independent experiments.
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fig02: Schematic representations of transcriptome responses in 3 day biofilm, 4 day biofilm and stationary-phase conditions. A. Conditions corresponding to the 3 day biofilm (green circle), 4 day biofilm (blue circle) and stationary phase (red circle) are shown, with the numbers of induced genes (> 4-fold) shown in black and the number of repressed genes (> 4-fold) shown in red. The numbers of induced and regulated genes common to two or more conditions are included at the intersections of the three circles. A complete list of microarray data for genes in each category is included in Table S2. B. Induction of genes involved in iron acquisition during biofilm and stationary-phase growth. Twenty-nine genes arranged in nine operons (Msmeg0011–0014, Msmeg0015, Msmeg0016–0018, Msmeg2133–2135, Msmeg4502–4509, Msmeg4510, Msmeg5028, Msmeg6024–6026 and Msmeg6214–6216) putatively involved in iron acquisition show substantial levels of induction after 4 days of biofilm development. Arrows indicate the operon arrangements. Average fluorescence units are shown for the exponential planktonic (blue bars), 3 day biofilm (red bars), 4 day biofilm (yellow bars) and stationary phase (green bars). Fluorescence values for the exponential planktonic sample are the average of 12 normalized slides, and for the other three experimental conditions, are the average from three independent experiments.

Mentions: The first question we wished to address was whether there were global changes in gene expression during M. smegmatis biofilm formation. We observe that there are relatively modest changes in the 3 day biofilm sample, with only ∼1.5% genes changing expression by four-fold or more (see Fig. S2). This pattern changes substantially by day 4 of biofilm growth, where ∼4.5% of the genome changes by fourfold for more (Fig. S2); a similar proportion of genes (approximately 4.9% of the genome) change expression in stationary phase (Fig. S2). In general, the majority of genes in the stationary-phase sample are repressed, while those in the biofilm samples are induced; many genes respond to two or more of the conditions tested. An illustration of these patterns is shown in Fig. 2A, and a list of genes is provided in Table S3.


The role of iron in Mycobacterium smegmatis biofilm formation: the exochelin siderophore is essential in limiting iron conditions for biofilm formation but not for planktonic growth.

Ojha A, Hatfull GF - Mol. Microbiol. (2007)

Schematic representations of transcriptome responses in 3 day biofilm, 4 day biofilm and stationary-phase conditions. A. Conditions corresponding to the 3 day biofilm (green circle), 4 day biofilm (blue circle) and stationary phase (red circle) are shown, with the numbers of induced genes (> 4-fold) shown in black and the number of repressed genes (> 4-fold) shown in red. The numbers of induced and regulated genes common to two or more conditions are included at the intersections of the three circles. A complete list of microarray data for genes in each category is included in Table S2. B. Induction of genes involved in iron acquisition during biofilm and stationary-phase growth. Twenty-nine genes arranged in nine operons (Msmeg0011–0014, Msmeg0015, Msmeg0016–0018, Msmeg2133–2135, Msmeg4502–4509, Msmeg4510, Msmeg5028, Msmeg6024–6026 and Msmeg6214–6216) putatively involved in iron acquisition show substantial levels of induction after 4 days of biofilm development. Arrows indicate the operon arrangements. Average fluorescence units are shown for the exponential planktonic (blue bars), 3 day biofilm (red bars), 4 day biofilm (yellow bars) and stationary phase (green bars). Fluorescence values for the exponential planktonic sample are the average of 12 normalized slides, and for the other three experimental conditions, are the average from three independent experiments.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2170428&req=5

fig02: Schematic representations of transcriptome responses in 3 day biofilm, 4 day biofilm and stationary-phase conditions. A. Conditions corresponding to the 3 day biofilm (green circle), 4 day biofilm (blue circle) and stationary phase (red circle) are shown, with the numbers of induced genes (> 4-fold) shown in black and the number of repressed genes (> 4-fold) shown in red. The numbers of induced and regulated genes common to two or more conditions are included at the intersections of the three circles. A complete list of microarray data for genes in each category is included in Table S2. B. Induction of genes involved in iron acquisition during biofilm and stationary-phase growth. Twenty-nine genes arranged in nine operons (Msmeg0011–0014, Msmeg0015, Msmeg0016–0018, Msmeg2133–2135, Msmeg4502–4509, Msmeg4510, Msmeg5028, Msmeg6024–6026 and Msmeg6214–6216) putatively involved in iron acquisition show substantial levels of induction after 4 days of biofilm development. Arrows indicate the operon arrangements. Average fluorescence units are shown for the exponential planktonic (blue bars), 3 day biofilm (red bars), 4 day biofilm (yellow bars) and stationary phase (green bars). Fluorescence values for the exponential planktonic sample are the average of 12 normalized slides, and for the other three experimental conditions, are the average from three independent experiments.
Mentions: The first question we wished to address was whether there were global changes in gene expression during M. smegmatis biofilm formation. We observe that there are relatively modest changes in the 3 day biofilm sample, with only ∼1.5% genes changing expression by four-fold or more (see Fig. S2). This pattern changes substantially by day 4 of biofilm growth, where ∼4.5% of the genome changes by fourfold for more (Fig. S2); a similar proportion of genes (approximately 4.9% of the genome) change expression in stationary phase (Fig. S2). In general, the majority of genes in the stationary-phase sample are repressed, while those in the biofilm samples are induced; many genes respond to two or more of the conditions tested. An illustration of these patterns is shown in Fig. 2A, and a list of genes is provided in Table S3.

Bottom Line: In contrast, although the expression of mycobactin and iron ABC transport operons is highly upregulated during biofilm formation, mutants in these systems form normal biofilms in low-iron (2 microM) conditions.A close correlation between iron availability and matrix-associated fatty acids implies a possible metabolic role in the late stages of biofilm maturation, in addition to the early regulatory role.M. smegmatis surface motility is similarly dependent on iron availability, requiring both supplemental iron and the exochelin pathway to acquire it.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260, USA.

ABSTRACT
Many species of mycobacteria form structured biofilm communities at liquid-air interfaces and on solid surfaces. Full development of Mycobacterium smegmatis biofilms requires addition of supplemental iron above 1 microM ferrous sulphate, although addition of iron is not needed for planktonic growth. Microarray analysis of the M. smegmatis transcriptome shows that iron-responsive genes - especially those involved in siderophore synthesis and iron uptake - are strongly induced during biofilm formation reflecting a response to iron deprivation, even when 2 microM iron is present. The acquisition of iron under these conditions is specifically dependent on the exochelin synthesis and uptake pathways, and the strong defect of an iron-exochelin uptake mutant suggests a regulatory role of iron in the transition to biofilm growth. In contrast, although the expression of mycobactin and iron ABC transport operons is highly upregulated during biofilm formation, mutants in these systems form normal biofilms in low-iron (2 microM) conditions. A close correlation between iron availability and matrix-associated fatty acids implies a possible metabolic role in the late stages of biofilm maturation, in addition to the early regulatory role. M. smegmatis surface motility is similarly dependent on iron availability, requiring both supplemental iron and the exochelin pathway to acquire it.

Show MeSH
Related in: MedlinePlus